When actually applied to the production of a certain desired substance by microorganisms, the recombinant DNA technology generally presents a problem that the recombinant plasmid is unstable.
The cloning and expression vectors generally used in laboratories are mostly multicopy plasmids, and the stable transmission thereof to later generations can be secured by introduction of a number of plasmids per cellular genome (cf. Jones I M et al., Mol. Gen. Genet. 180(3): 579-84 (1980)). However, when a foreign gene is introduced using a plasmid, plasmid elimination occurs during bacterial proliferation cycles, resulting in the instability of the gene introduced. Therefore, in a commercial production process, it is essential to stabilize the plasmid in bacteria until completion of the culture in a fermentor.
So far pJRD215-derived vectors and pBBR-derived vectors have been mainly used as plasmid vectors for gene transfer into bacteria of the genus Ralstonia, Wautersia and Cupriavidus (T. Fukui et al., Biotechnology Letters, Vol. 19, No. 11, November 1997; 1093-97); and Steven Slater et al., Journal of Bacteriology, April 1998: 1979-1987). However, investigations made by the present inventors have revealed that these plasmid vectors become very unstable upon accumulation of polyhydroxyalkanoates, among others, in cells by host bacteria. In the case of a pJRD215-derived vector, for instance, about 80% of the cells after 4 passages under cultivation conditions under which the accumulation of polyhydroxyalkanoates is not rich, without application of any antibiotic-due selective pressure, retain the plasmid, whereas, after 4 passages under cultivation similarly conditions allowing a great accumulation of the polyesters, only 30% of the cells retain the plasmid. The same characteristic feature is presented by pBBR-derived vectors.
These plasmid vectors are broad-host-range vectors developed for use in a wide range of hosts (cf. Luan Tao et al., Metabolic Engineering, Volume 7, Issue 1, January 2005: 10-17; and Davison J et al., Gene, 1987; 51(2-3): 275-80). However, in commercial substance productions using bacteria of the genus Ralstonia, Wautersia or Cupriavidus, for instance, as the hosts, it is necessary to develop plasmid vectors for gene transfer suited for use in bacteria of the genus Ralstonia, Wautersia or Cupriavidus. 
Various techniques have so far been developed to stabilize plasmids. However, as regards plasmid vectors capable of being used in the genus Ralstonia, it is only possible to select plasmid-retaining transformants according to the resistance to such a drug as chloramphenicol, kanamycin or ampicillin. When the cultivation is carried out under application of an antibiotic-due selective pressure, the following problems arise: (1) the use of an antibiotic-resistant strain may possibly involve risk to the environment, (2) the amount of an antibiotic which is required during cultivation significantly increases the cost of production, and (3) the use of antibiotics is undesirable for the production of substances to be used in the treatment of humans and animals. Thus, transformants retaining a plasmid owing to drug resistance cannot be applied to commercial production.
In addition to the technique comprising applying an antibiotic-due selective pressure, the so-called par system is known as a system for stable retention of a plasmid (cf. M. Gerlitz et al., Journal of Bacteriology, November: 6194-6203 (1990); and B. Youngren et al., Journal of Bacteriology, July: 3924-3928 (2000)). When the par system works, the plasmids replicated are partitioned among daughter cells, so that a bacterial strain stably retaining a plasmid can be obtained without applying any selective pressure due to antibiotic resistance. Thus, a vector with the par system of the RP4 plasmid, which is usable in Escherichia coli, inserted therein (cf. U.S. Pat. No. 6,143,518) and a vector in which the par region of the R1 plasmid is utilized (cf. U.S. Pat. No. 4,760,022) have so far been developed. Further, it is also known that, in the par region of the megaplasmid pMOL28 retained in the Cupriavidus metallidurans CH34 strain, there occur the promoter parP, the plasmid stabilizing factors parA28 and parB28 and the recognition sequence pars; these genes have already been cloned in the plasmid pSUP202 and the nucleotide sequences thereof have been published (cf. Safieh Taghavi et al., Mol. Gen. Genet. 250: 169-179 (1996)).
As described above, since bacteria belonging to the genus Ralstonia, Cupriavidus or Wautersia (in particular the species Cupriavidus necator) are often used as bacteria for the production of polyhydroxyalkanoates or the production of proteins (cf. Gravin C. et al., Protein Expression & Purification December; 38(2): 64-71 (2004)), it is required to develop a plasmid vector which is particularly usable in these bacteria, has no transferability by conjugation, and is capable of being stably retained therein without any antibiotic-due selective pressure. However, as far as bacteria belonging to the genus Ralstonia, Cupriavidus or Wautersia are concerned, such plasmid stabilization using the par region has not been made as yet; how to construct such system or whether such system is actually effective for plasmid stabilization is unknown.